Proteomics of malignant pleural mesothelioma under hypoxic and normoxic conditions in a large animal (porcine) tumor model

Malignant Pleural Mesothelioma (MPM) is a rare but highly aggressive malignant tumor, whose occurrence is closely associated with asbestos exposure. Clinical treatment options are limited and the prognosis is poor. The tumor microenvironment, especially hypoxia, plays a key regulatory role in the occurrence and development of MPM, yet its specific molecular mechanism remains unclear. To explore the effects of different oxygen metabolic states on the protein expression profiles and functional characteristics of MPM, this study first cultured MPM cells under normoxic and hypoxic conditions and conducted proteomic analysis. Furthermore, it explored MPM animal models simulating normoxic and hypoxic conditions, aiming to provide an experimental platform closer to the physiological state for MPM mechanism research and targeted intervention. In in vitro studies, MSTO-211 H cells were cultured under 1% O₂ concentration (experimental group) and 21% O₂ concentration (control group) for 48 h, passaged for 3 generations, with 3 biological replicates in each group. TMT quantitative proteomics was used to screen differential proteins, and GO and KEGG analyses were performed to explore related signaling pathways. In animal experiments, the hypoxic group and normoxic group were injected with benzo(a)pyrene solution via a closed thoracic device, while the blank control group was injected with normal saline. Clinical symptoms and multiple biological indicators of the animals were monitored, and tumor characteristics were described based on CT, histopathological and immunohistochemical results.​ In in vitro studies, a total of 42 differential proteins were identified between the hypoxic group and the normoxic group. GO and KEGG analyses revealed that the two groups shared differences in three signaling pathways: protein digestion and absorption, neuroactive ligand-receptor interaction, and Staphylococcus aureus infection. In in vivo studies, MPM occurred in both the hypoxic group (Tibetan pigs) and the normoxic group (Landrace pigs) after intrathoracic injection of benzo(a)pyrene, while the normoxic group was accompanied by benign pleural hyperplasia. Only inflammatory cell infiltration was observed in the normal saline control group. The tumor tissue was composed of well-differentiated epithelioid cells and fibrovascular stroma. IHC detection showed that the tumor cells expressed specific proteins consistent with MPM, and the tumor microenvironment contained abundant CD31-positive small vascular endothelial cells. During the follow-up period of 224–238 days, the experimental pigs remained in good health.​ Based on the study of differentially expressed proteins in malignant pleural mesothelioma under hypoxic and normoxic conditions, the successfully constructed large-scale porcine tumor model can pre-verify the effect of drugs on tumors in different oxygen environments during new drug development, improving screening efficiency and success rate. It can also accurately simulate clinical scenarios, providing experimental support for the efficacy evaluation and scheme optimization of intervention strategies such as chemotherapy, radiotherapy, and immune targeting, thereby effectively promoting the development of clinical treatment for malignant pleural mesothelioma.​